The focus of this proposal is to study the basic mechanisms underlying axonopathy and progressive neurodegeneration that are caused by mutations in the swiss cheese (sws) gene. SWS is the ortholog of human Neuropathy Target Esterase (NTE), a key player in organophosphate (OP)-induced delayed neuropathy (OPIDN) which is caused by organophosphates found in many pesticides and nerve agents. Neuronal loss of NTE in mice results in a pattern of progressive degeneration of the nervous system that is surprisingly similar to mutant flies, while the complete loss of NTE is embryonic lethal. In addition, recently described mutations in the human gene cause a hereditary spastic paraplegia called NTE--related Motor Neuron Disorder. The functional conservation between SWS and NTE was confirmed by experiments showing that mouse NTE can replace the fly protein, reverting the degenerative phenotype caused by the loss of SWS. SWS and NTE can both hydrolyze the same artificial substrate, and both have been connected with membrane lipid homeostasis; however, their endogenous functions are still largely unknown. During the previous funding period, several domains in the SWS protein were identified and shown to be functionally required. This includes a domain that binds a specific catalytic subunit of protein kinase A (PKA-C3), thereby inhibiting its kinase activity. PKA-C3, and its vertebrate orthologs PrKX in human and Pkare in mice, form a novel class of catalytic subunits that are more closely related to each other than to other catalytic subunits within the same species. They are all expressed in the nervous system, but their functions are so far unknown. The current proposal will tests the hypothesis that SWS and PKA-C3 form an unconventional PKA complex that has a dual function: regulating the intracellular localization and kinase activity of PKA-C3, and controlling the esterase activity of SWS. Furthermore, the role of this unique PKA subunit in neural survival, sws-induced phenotypes, and organophosphate induced toxicity will be investigated. Due to the high conservation of these proteins between Drosophila and vertebrates, the proposed studies in this well-established model system can provide important insights into the mechanisms that lead to progressive degeneration in the related human disease and in toxin-induced neuropathy. Beyond the scope of the immediate proposal, these studies may also expand our understanding of common mechanisms of axonopathy and neuronal cell death, which also occurs in response to a variety of other neuronal insults.